Simian virus 40-specific proteins in HeLa cells infected with nondefective adenovirus 2-simian virus 40 hybrid viruses.

The constitutive transcription of a mouse alpha-fetoprotein (AFP) minigene was examined during the transient expression of AFP-simian virus 40-pBR322 recombinant DNAs introduced into HeLa cells by Ca3(PO4)2 precipitation. We tested three constructs, each of which contains the AFP minigene and pBR322 DNAs inserted in the late region of simian virus 40 and found that the relative efficiency of AFP gene expression was dependent on the arrangement of the three DNA elements in the vector. The transcripts begin at the authentic AFP cap site and are properly spliced and polyadenylated. To define a sequence domain in the 5' flanking region of the AFP gene required for constitutive expression, sequential 5' deletion mutants of the AFP minigene were constructed and introduced into HeLa cells. All AFP deletion mutants which retained at least the TATA motif located 30 base pairs upstream from the cap site were capable of directing accurate and efficient AFP transcription. However, when the TATA sequence was deleted, no accurately initiated AFP transcripts were detected. These results are identical to those obtained from in vitro transcription of truncated AFP 5' deletion mutant templates assayed in HeLa cell extracts. The rate of AFP transcription in vivo was unaffected by deletion of DNA upstream of the AFP TATA box but was greatly affected by the distance between the simian virus 40 control region and the 5' end of the gene. The absence of any promoter activity upstream of the TATA box in this assay system is in contrast to what has been reported for several other eucaryotic structural genes in a variety of in vivo systems. A sequence comparison between the 5' flanking region of the AFP gene and these genes suggested that the AFP gene lacks those structural elements found to be important for constitutive transcription in vivo. Either the AFP gene lacks upstream promoter function in the 5' flanking DNA contained within the minigene, or the use of a viral vector in a heterologous system precludes its identification.

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Transcriptional control of the mouse prealbumin (transthyretin) gene: both promoter sequences and a distinct enhancer are cell specific.

Molecular and Cellular Biology ◽

10.1128/mcb.6.12.4697 ◽

1986 ◽

Vol 6 (12) ◽

pp. 4697-4708 ◽

Cited By ~ 77

Author(s):

R H Costa ◽

E Lai ◽

J E Darnell

Keyword(s):

Hela Cells ◽

Hepg2 Cells ◽

Simian Virus 40 ◽

Simian Virus ◽

Proximal Promoter ◽

Start Site ◽

Beta Globin ◽

Human Hepatoma Cells ◽

Enhancer Element ◽

A Cell

The mouse genomic clone for the prealbumin (transthyretin) gene was cloned, and its upstream regulatory regions were analyzed. The 200 nucleotides 5' to the cap site when placed within a recombinant plasmid were sufficient to direct transient expression in HepG2 (human hepatoma) cells, but this DNA region did not support expression in HeLa cells. The sequence of the 200-nucleotide region is highly conserved between mouse and human DNA and can be considered a cell-specific promoter. Deletions of this promoter region identified a crucial element for cell-specific expression between 151 and 110 nucleotides 5' to the RNA start site. A region situated at about 1.6 to 2.15 kilobases upstream of the RNA start site was found to stimulate expression 10-fold in HepG2 cells but not in HeLa cells. This far upstream element was invertible and increased expression from the beta-globin promoter in HepG2 cells. Unlike the simian virus 40 enhancer, the prealbumin enhancer would not stimulate beta-globin synthesis in HeLa cells, and even the simian virus 40 enhancer did not stimulate the prealbumin promoter in HeLa cells. Thus, we identified in the prealbumin gene two DNA elements that respond in a cell-specific manner: a proximal promoter including a crucial sequence between -108 and -151 nucleotides and a distant enhancer element located between 1.6 and 2.15 kilobases upstream.

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Structural relationship between the 100,000- and 17,000- molecular-weight T antigens of simian virus 40 (SV40) as deduced by comparison with the SV40-specific proteins coded by the nondefective adenovirus type 2-SV40 hybrid viruses.

Journal of Virology ◽

10.1128/jvi.29.1.390-394.1979 ◽

1979 ◽

Vol 29 (1) ◽

pp. 390-394 ◽

Cited By ~ 10

Author(s):

H K Linke ◽

T Hunter ◽

G Walter

Keyword(s):

Molecular Weight ◽

Simian Virus 40 ◽

Adenovirus Type ◽

Simian Virus ◽

Structural Relationship ◽

T Antigens ◽

Specific Proteins ◽

Adenovirus Type 2

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Suppression of tropomyosin synthesis, a common biochemical feature of oncogenesis by structurally diverse retroviral oncogenes

Molecular and Cellular Biology ◽

10.1128/mcb.5.5.972-983.1985 ◽

1985 ◽

Vol 5 (5) ◽

pp. 972-983

Author(s):

H L Cooper ◽

N Feuerstein ◽

M Noda ◽

R H Bassin

Keyword(s):

Cell Lines ◽

Simian Virus 40 ◽

Simian Virus ◽

Transformation Process ◽

Cytoskeletal Proteins ◽

Morphological Alteration ◽

Initial Appearance ◽

Specific Proteins ◽

3T3 Cell Lines ◽

Nih 3T3

To identify proteins whose production may be altered as a common event in the expression of structurally diverse oncogenes, we compared two-dimensional electropherograms of newly synthesized proteins from NIH/3T3 cell lines transformed by a variety of retroviral oncogenes, from cellular revertant lines, and from a line (433.3) which expresses the v-ras oncogene in response to corticosteroids. Most alterations in the synthesis of specific proteins detected by this approach appeared to be the result of selection during prolonged cultivation and were probably unrelated to the transformation process. However, we detected seven proteins whose synthesis was strongly suppressed in cell lines transformed by each of the six retroviral oncogenes we studied and whose production was fully or partially restored in two cellular revertant lines. Suppression of two of these proteins was also correlated with the initial appearance of morphological alteration during corticosteroid-induced oncogene expression in 433.3 cells. These proteins (p37/4.78 and p41/4.75) were identified as tropomyosins, a group of at least five cytoskeletal proteins. Transformation by the papovaviruses simian virus 40 and polyomavirus caused no suppression of synthesis of these tropomyosins. This indicates that suppression of tropomyosin synthesis is not a nonspecific response by cells to being forced to grow with the transformed phenotype but is specifically associated with oncogenesis by diverse retroviral oncogenes. The results are consistent with the hypothesis that the different biochemical processes initiated by expression of structurally diverse retroviral oncogenes may converge on a limited number of common targets, one of which is the mechanism which regulates the synthesis of tropomyosins.

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